Univ. of Wisconsin School of Medicine and Public Health, 5229 MFCB 1685 Highland Ave, Madison, WI 53705, USA.
Am J Physiol Lung Cell Mol Physiol. 2011 Nov;301(5):L656-66. doi: 10.1152/ajplung.00166.2011. Epub 2011 Aug 19.
Myofibroblast differentiation induced by transforming growth factor-β (TGF-β) and characterized by de novo expression of smooth muscle (SM)-specific proteins is a key process in wound healing and in the pathogenesis of fibrosis. We have previously shown that TGF-β-induced expression and activation of serum response factor (SRF) is required for this process. In this study, we examined the signaling mechanism for SRF activation by TGF-β as it relates to pulmonary myofibroblast differentiation. TGF-β stimulated a profound, but delayed (18-24 h), activation of Rho kinase and formation of actin stress fibers, which paralleled SM α-actin expression. The translational inhibitor cycloheximide blocked these processes without affecting Smad-dependent gene transcription. Inhibition of Rho kinase by Y-27632 or depolymerization of actin by latrunculin B resulted in inhibition TGF-β-induced SRF activation and SM α-actin expression, having no effect on Smad signaling. Conversely, stabilization of actin stress fibers by jasplakinolide was sufficient to drive these processes in the absence of TGF-β. TGF-β promoted a delayed nuclear accumulation of the SRF coactivator megakaryoblastic leukemia-1 (MKL1)/myocardin-related transcription factor-A, which was inhibited by latrunculin B. Furthermore, TGF-β also induced MKL1 expression, which was inhibited by latrunculin B, by SRF inhibitor CCG-1423, or by SRF knockdown. Together, these data suggest a triphasic model for myofibroblast differentiation in response to TGF-β that involves 1) initial Smad-dependent expression of intermediate signaling molecules driving Rho activation and stress fiber formation, 2) nuclear accumulation of MKL1 and activation of SRF as a result of actin polymerization, and 3) SRF-dependent expression of MKL1, driving further myofibroblast differentiation.
转化生长因子-β（TGF-β）诱导的肌成纤维细胞分化，其特征是平滑肌（SM）特异性蛋白的从头表达，是伤口愈合和纤维化发病机制中的关键过程。我们之前已经表明，TGF-β诱导的血清反应因子（SRF）的表达和激活对于这个过程是必需的。在这项研究中，我们研究了 TGF-β激活 SRF 的信号机制，因为它与肺肌成纤维细胞分化有关。TGF-β刺激了 Rho 激酶的深刻但延迟（18-24 小时）激活和肌动蛋白应力纤维的形成，这与 SM α-肌动蛋白的表达平行。翻译抑制剂环己酰亚胺阻断了这些过程，而不影响 Smad 依赖性基因转录。Rho 激酶抑制剂 Y-27632 或肌动蛋白解聚剂 latrunculin B 的抑制作用导致 TGF-β诱导的 SRF 激活和 SM α-肌动蛋白表达的抑制，对 Smad 信号没有影响。相反，jasplakinolide 稳定肌动蛋白应力纤维足以在没有 TGF-β的情况下驱动这些过程。TGF-β促进了 SRF 共激活因子巨核细胞白血病-1（MKL1）/心肌相关转录因子-A 的延迟核积累，而 latrunculin B 抑制了这一过程。此外，TGF-β还诱导了 MKL1 的表达，latrunculin B、SRF 抑制剂 CCG-1423 或 SRF 敲低均可抑制 MKL1 的表达。综上所述，这些数据表明，肌成纤维细胞对 TGF-β的反应存在一个三相模型，包括 1）最初的 Smad 依赖性中间信号分子表达，驱动 Rho 激活和应力纤维形成，2）由于肌动蛋白聚合导致 MKL1 的核积累和 SRF 的激活，以及 3）SRF 依赖性 MKL1 的表达，驱动进一步的肌成纤维细胞分化。
